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Purification and Characterization of β‐Glucosidase from Brassica oleracea

Author:
Bešić, Larisa, Ašić, Adna, Muhović, Imer, Dogan, Serkan, Turan, Yusuf
Source:
Journal of food processing and preservation 2017 v.41 no.2
ISSN:
0145-8892
Subject:
Brassica oleracea, ammonium sulfate, beta-glucosidase, broccoli, enzyme activity, gluconolactone, glucose, hydrophobic interaction chromatography, industrial applications, pH, salting, temperature
Abstract:
β‐Glucosidase was purified from Brassica oleracea by salting out with ammonium sulfate and hydrophobic interaction chromatography. Results demonstrated that the enzyme is a dimer (130 kD) made up of one major (80 kD) and one minor subunit (50 kD). The pH optimum is 6.0, with 50% of the enzyme's original activity remaining between pH 4.0 and pH 7.0. The temperature optimum is 35C, and activity did not decrease after two hours of exposure to this temperature. The activity of the enzyme was investigated on four substrates, 4‐Nitrophenyl β‐D‐glucopyranoside (p‐NPG), ortho‐Nitrophenyl‐β‐D‐glucopyranoside (o‐NPG), para‐Nitrophenyl‐β‐D‐galactoside (p‐NPGal) and ortho‐Nitrophenyl‐β‐D‐galactoside (o‐NPGal), and kₘ values were shown to be 0.755 mM, 0.174 mM, 0.988 mM and 0.213 mM, while Vₘₐₓ values were 604 U/mg, 38 U/mg, 556 U/mg and 308 U/mg, respectively. The enzyme is completely inhibited by gluconolactone and glucose against p‐NPG as substrate, with kᵢ values of 0.038 mM and 0.64 mM, respectively. To our knowledge, this is the first study demonstrating purification and characterization of β‐glucosidase from broccoli, thus providing a better understanding of its role in the plant, and establishing a basis for further research. PRACTICAL APPLICATIONS: To our knowledge, this is the first study demonstrating purification and characterization of β‐glucosidase from broccoli, thus providing a better understanding of its role in the plant, and establishing a basis for further research. The results of this research highlight the potential of the enzyme isolated from broccoli for further research. Succeeding efforts would involve optimization of this procedure for increasing the enzyme yield, in order to make it a viable candidate for industrial application.